Journal of General Microbiology (1984), 130, 983-990.
Printed in Great Britain
983
Effects of Sedimentation and Light Intensity on
Mat-forming Oscillatoviaceae with Particular Reference to
Microcoleus iyngbyaceus Gomont
By A L L A N P E N T E C O S T
Department of Biological Sciences, Chelsea College, University of London,
London SWIO OQX,UK
(Received 27 October 1983)
The distribution and abundance of mat-forming UsciZlatoriaceae were investigated at a siltdepositing freshwater site in England over a period of 4 years. Maximum growth occurred
during April and May and experiments showed that mat buried by fine sediment could regain
the surface by gliding movements. The number of trichomes reaching the surface was lightdependent but chemotactic responses were also apparent. Gliding rates of 1-2 ym s- were
recorded and the trichomes moved through the sediment as a left-handed screw with a pitch of
60". A surface illumination of 1 klx (4W m - ') was optimal for mat formation. Light intensities
exceeding 2-5 klx produced a photophobic response resulting in lateral trichome migration to
regions of lower surface illumination when these were provided. Migration into the sediment
was not observed in the surface intensity range 0-10 klx. The onset of light saturation measured
by the 14C02 method was 3 klx for Microcoleus lyngbyaceus, close to the optimum for mat
development. Experiments employing l4COZin the presence of sulphide and absence of oxygen
suggested that the organisms were adapted to an aerobic environment.
INTRODUCTION
Mat-forming cyanobacteria belonging to the family Oscillatoriaceae cover extensive areas of
shallow water subject to sedimentation throughout the world. In some regions, usually where the
water is saline or depositing carbonates, the mats have a considerable stabilizing effect and may
bind the sediments sufficiently well to produce a durable substratum (Golubic, 1973; Walter,
1976; Pentecost, 1978) whilst in other sites there is little binding, the development of mats is
seasonal and the sediment is shifting continuously. This last environment, although of worldwide occurrence, has received little attention and has not been investigated experimentally. An
accreting benthos would appear to hold few advantages for a photoautotrophic organism since
survival would depend upon an efficient means of overcoming the obstruction of light caused by
the rain of sediment. The ability to perform net photosynthesis under low light regimes
associated with turbid water would also be of advantage. Previous studies would suggest that
members of the Oscillatoriaceae have the potential to overcome these disadvantages as many are
motile and some planktonic forms are saturated for photosynthesis at low light levels
(Zimmermann, 1969). However, data relating to motility in natural habitats are primarily
qualitative (Castenholz, 1982) and there are no quantitative estimates of growth of these forms
on sediment surfaces. This investigation attempts to describe the cyanobacteria associated with
a temperate silt-depositing site and to determine their ability to withstand rapid sedimentation
under differing light regimes.
METHODS
Sampling and characterization of cyanobacteria. Samples were collected from a small tributary of the River
Medway near Tonbridge, Kent, UK (NGR 51/577465) using a modification of the method described by Round
& Heaton (1966). The suspension of silt and cyanobacteria was allowed to settle for 5 h and the total trichome
0022-1287/84/0001-1298 $02.00 0 1984 SGM
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A . PENTECOST
length measured under a dissection microscope ( x 50) under a surface illumination of 2 klx. Trichome length was
calculated by the grid method described by Olson (1950). Since the trichomes were initially Poisson distributed on
the sediment surface, confidence limits could be calculated for the length estimates. The occurrence of sheath
material and polyphosphate bodies was monitored using ruthenium red and Ebel's solution respectively (Fuhs,
1973). Rates of motility were recorded under the microscope at 20°C (illumination 5001x) with a stopwatch.
Trichomes were photographed using Ilford FP4 film and width measurements were made under oil immersion
with a Leitz Ortholux microscope. Site temperature, illumination and water chemistry were also monitored
(Mackereth et af., 1978).
Sedimentation studies. Silt samples from the site were well washed, sieved and autoclaved before use and a slurry
was prepared by decanting the supernatant water after standing for 3 h. Mats were allowed to develop in Petri
dishes with the sides covered in aluminium foil and a calculated volume of silt suspension was carefully poured
over the mat to give depths ranging from 2.5 to 7.5 mm. The surfaces were illuminated in the range 0-1-20 klx and
the temperature was maintained at 15°C. The number of trichomes appearing at the sediment surface was
measured at regular intervals as described above.
Incorporation ofH14CO;. Trichomes of Microcoleus fyngbyaceus were harvested free of silt by collecting them
from the sides of glass containers and brought into suspension by shaking with Chu 10 medium at pH 7.0 (Chu,
1942). Suspensions were incubated at 15 "C under light intensities of 0-1-20 klx with 10 pCi NaH14C03of specific
activity 3.3 mCi mmol-l (122 MBq mmol-l). To study the effect of deoxygenation, two suspensions were
prepared, one sparged with nitrogen and the other with air for 2 h prior to incubation. Suspensions were incubated
at light saturation in sealed 20 ml glass bottles. Another set of deoxygenated samples was spiked with up to 0.5 mMNazS at pH 7.0 and here the Chu 10 was modified to prevent sulphide precipitation by decreasing the
.
6 to 10 replicates were prepared of all treatments and after fixation in
concentration of Ca and Mg to 5 p ~From
1 % (v/v) formalin the samples were processed by bubbling and acidification (Schindler et a[., 1972). The samples
were then added to Aquasol scintillation fluor and counted on a Beckman LS 230 scintillation counter with an
efficiency of 78-80%. Uptake rates were corrected for dark uptake, although this was negligible. Illumination for
all experiments was provided by Thorn 'warm white' fluorescent tubes.
RESULTS
Distribution and characterization
Trichome width measurements were used as a basis of classification and a unimodal or bimodal distribution was noted (Fig. 1). The dominant form had a mode class width of 13-16 pm
and was identified as Microcoleus fyngbyaceus Gomont with olive-brown to blue-green trichomes
and rounded end cells (Fig. 2a). A narrow form with a mode class width of 4-6 pm was abundant
during 1981 (Fig. 1). It had hooked trichome ends and was identified as Osciffatoriabrevis
(Kutz.) Gomont (Fig. 2 b). An intermediate form occurred during 1980 with trichomes 11-12 pm
wide, resembling a large form of 0. brevis. The trichomes also had hooked ends.
Trichomes were found on the sediment surface throughout the year in small numbers but
maximum biomass was noted during April and May (Fig. 3a, b) after which a rapid decline
occurred. During April and May water temperature ranged from 4 to 12 "C but there was no
correlation between temperature and the numbers recorded. Maximum light intensity at the
water surface remained approximately constant throughout the year due to dense shading by
trees during the summer, but extreme variations were recorded at the sediment surface at noon
(<0.05-15 klx) depending upon the water turbidity. The length of individual trichomes ranged
from <0-05 to 2.4mm but usually averaged 0.5-1 mm in both species. Trichome length
appeared to be significantly greater early in the year (Fig. 3c, d ) during the development of
locally dense mats. The frequency distribution of trichome length in M . fyngbyaceus during May
showed an approximately log-normal form (Fig. 3 1 ) with large numbers of short hormogonia
present. A significant correlation (P< 0.05, Spearman's test) was found between trichome
length frequency and the frequency of the distance between successive necridia in intact
trichomes (Fig. 3 e). This suggests that the sampling procedure caused insignificant damage
through trichome breakage. The cells of both species possessed randomly distributed polyphosphate bodies throughout the year and firm, permanent sheaths such as those found in Ljmgb-va
were never observed.
Water at the sampling site was 5-40 cm deep and was eutrophic (pH 6.8-7.5, total COz
2.4-2-6 mM, Ca 3.5 mM, Mg 0.3 mM, total P 8-10 VM,NO, 0.1-0-25 mM, total sulphide <0.2 mM,
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Mat formation by Microcoleus
L
-uluu
15
20
20
0
5
10
15
Trichome diameter (pm)
20
Fig. 1. Frequency distributions of trichome widths for samples collected in May 1980-83.
Fig. 2. (a) Trichomes of M . lyngbyaceus showing discoid cells, necridia and rounded apices.
(b) Trichomes of 0.breuis (left) with rostrate apices, and two trichomes of M . lyngbyaceus (right).
Bars 25 pm.
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A . PENTECOST
50
0
J F M A M J J A S O N D
75
50
25
0
Distance between necridia (mm)
0.5
1.0
Trichome length (mm)
>1+3
Fig. 3. (a, b) Mean trichome biomass, expressed as trichome length (cm) per cm2 of sediment for 0.
brevis ( a ) and M . lyngbyaceus (b).(c, d ) Mean trichome length for 0. brevis (c) and M . lyngbyaceus ( d )
during 1981. (e) Frequency distribution of the distances between adjacent necridia in long trichomes of
M . lyngbyaceus. (s)Frequency distribution of trichome length in the same sample.
oxygen-saturated). The sediment consisted of fine silt of mean diameter 13.5 pm and range
<0.5-75 pm with about 4% organic matter. Eh and pH of the sediment surface ranged from
+ 190 to +430 mV and from 6.8 to 7.5 respectively. There was a black anoxic zone at 2-3 cm
depth and the light extinction coefficient for the sediment was 18 mm- l .
Response of M . lyngbyaceus to sedimentation
Burial beneath 2.5 mm sediment resulted in almost complete recovery of mats within 3 h with
a surface light intensity of 2klx (Fig. 4 a ) . Burial under 5mm gave a similar initial rate of
recovery but this was not sustained. Burial beneath 7.5 mm sediment resulted in the emergence
of only 30% of the mat after 5 h (Fig. 4a). The recovery rate in the light was similar to that in the
dark under 5 mm sediment. Comparable results were obtained with 0 . brevis.
Trichome emergence was influenced by the light intensity with the greatest recovery
occurring at 1 klx when mats were buried beneath 3 mm sediment (Fig. 4 b). The same result was
obtained when recovery was recorded after 8 h burial using a greater range of surface light
intensities (Fig. 4c). In both experiments, recovery in the dark was significant but always lower
than that obtained with the optimum surface illumination.
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987
Mat formation by Microcoleus
100
100
n
8
W
x
0
$ 50
s
50
2
d
0
=
0
2
Time (h)
0
6
4
-1
0
+1
log,, (Light intensity, klx)
100
h
1
0.5
-8
2
Time (h)
3
1.0
2.5
5.0
Light intensity (klx)
10.0
5
10
15
Light intensity (klx)
20
100
v
8
c
v
.-
Y
(D
(c"
0
2
4
Time (h)
6
8
50
Fig. 4. (a) Recovery of M . lyngbyaceus after burial beneath sediment of the following depths: 2.5 mm
(0);
5 mm ( 0 ) 7.5
; mm
B, Dark control, 5 mm. Trichome length per unit area was measured at
various times after burial, and expressed as a percentage of the value before burial. (b)Recovery after
burial beneath 3 mm sediment as a function of surface light intensity: 1 klx (0);
5 klx ( 0 ) ;10 klx (U);
dark (B). (c) Recovery after 8 h burial beneath 3 mm sediment as a function of light intensity (log
scale). (d)Percentage of trichomes remaining on the illuminated half of a mat after 24h. The other half
was shaded by foil. Total trichomes on entire mat = 100%. (e) Recovery of mats buried beneath 3 mm
sediment with illumination at 1 klx. Light intensity was increased to 10 klx (arrow, 0)
after 4 h. U,
Control at 1 klx. (f)Relative photosynthetic 14C02uptake as a function of light intensity. The vertical
bars in (a), (b), (c) (e) and (f)represent 95% confidence limits.
(a).
Two further experiments were conducted to test for trichome migration away from the surface. When part of a mat was shaded by foil, trichomes were only found to migrate into light of
intensity 1 klx. At intensities of 0.5, 2.5 and 5 klx there was little or no tendency to migrate in
either direction but at 10 klx, trichome migration away from the light was significant (Fig. 4d).
This migration was lateral: no vertical migration was detected. When a mat was illuminated
initially at 1 klx and the illumination then increased to lOklx, vertical migration of the
trichomes was again found to be insignificant over a period of 4 h (Fig. 4e).
The mean gliding rates of M . lyngbyaceus and 0.breuis were 2-1 (range 1.95-2.2) and 1.33
(range 1.08-1.58) pm s - respectively at 20 "C and 0.5 klx. Both species moved through the
sediment in the sense of a left-handed screw with a pitch of 59-63". A significant negative
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A . PENTECOST
correlation (r = -0.78, P < 0.05) was found between trichome width and helix pitch in 0.breuis
but there was no correlation between the gliding rate and trichome length,
The rate of mat recovery was found to be partially dependent upon the nature of the sediment,
since mats covered with 3 mm clean, washed sand of 360 pm average diameter failed to recover
after 6 h although the trichomes remained motile.
Uptake of I4CO2
The relationship between 14C02uptake and light intensity is shown in Fig. 4 0 . The onset of
light saturation occurred at 3 klx and above 5 klx a decline in uptake was evident. When
trichomes were incubated with sulphide adjusted to pH 7, uptake showed a continuous decline
as sulphide concentrations increased from 0.02 to 0.5 mM and was almost entirely inhibited at
0.5 mM when compared with a deoxygenated control. When aerated and deoxygenated uptake
rates were compared, the former were found to be significantly higher than the latter (P = 0.05).
DISCUSSION
The sedimentation rates calculated for the site investigated, and the depth of sediment likely
to cover the trichomes, were less than the maximum employed in the experiments. This suggests
that mats are capable of re-forming at the surface even in the absence of light. This would be
important to the organisms since light intensities at the sediment surface were extremely low
when sedimentation was in process.
The inhibition of l4CO, uptake under anoxic conditions and in the presence of sulphide suggests that gradients of oxygen and sulphide within the sediment might be the stimulus causing
the trichomes to migrate in the dark. The occurrence of chemotactic behaviour is also supported
by the observation that trichomes moved away from the sediment surface in the dark when an
oxygen gradient was present. Castenholz (1982) noted a chemotactic response to sulphide by
Oscillatoria terebrformis, but there are problems in identifying the response since high sulphide
levels are usually associated with an absence of oxygen and elevated levels of ferrous iron and
carbon dioxide, any one of which might be the causative factor. The rate of recovery of
trichomes in the dark might be expected to fall as the depth of sediment above is increased. This
is apparent from the results (Figs. 4 a , b) but the effect is not so great as might be expected. The
cause of this is unclear but may be due to differences in concentration of dissolved substances in
different batches of sediment or variation in the respiratory activity of the buried trichomes,
which must affect the immediate environment.
With an average gliding rate of 2.1 pm s - l , M . lyngbyaceus should be capable of surfacing
within 2 h when buried beneath 7-5mm of silt, but the rate of recovery was much lower than this.
One possible explanation is a reduction in motility caused by the increased sediment compaction
with depth, but it is more likely to be a result of the longer path length the trichomes took to
reach the surface. The trichomes were unable to orientate themselves perpendicular to the surface, and arrived at low angles, usually less than lo", so that the distance travelled was much
greater. This also helps to explain the incomplete recovery of deeply buried mats. The trichomes
were found to move along straight paths through the sediment since the particles were small and
easily pushed aside. The inability of trichomes to move through sand appears to result from the
fact that movement in straight lines was excluded and the efficient screwing-motion made
ineffective.
Light-induced trichome motility has been investigated in detail in the laboratory. Optimum
photokinesis in Phormidium ambiguum occurred at 1 klx (Nultsch & Hellman, 1972) and at
0.2 klx in P.autumnale (Nultsch, 1962). The cyanobacteria so far investigated exhibit positive
phototaxis in the range 0 4 - 1 0 klx (Castenholz, 1982) so it would seem that the group as a
whole, with the Oscillatoriaceae in particular, are adapted well to low light regimes. Light
intensity is clearly the main factor controlling mat formation in M . lyngbyaceus and the response
of trichomes to different light regimes is of considerable interest. It would appear from these
experiments that trichomes, once at the sediment surface, prefer to glide laterally rather than
back into the sediment when the light regime becomes unsuitable. This behaviour may be due to
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Mat .formation by Microcoleus
989
the presence of substances dissolved in the interstitial water which produce a negative chemotactic response, but this conflicts with the results where the trichomes remained buried yet
apparently viable and motile. The trichomes are apparently incapable of migrating into the
sediment and can only alter their light regime by lateral migration, by moving over each other
and causing self-shading or by awaiting further sedimentation. Migration back into the
sediment is unlikely to provide a particularly suitable light climate for the trichomes unless fine
adjustments of position could be made. For example, with a surface illumination of 15 klx, the
trichomes would need to be positioned beneath 0.1 1-0.18 mm of sediment to achieve the
optimum light intensity of around 0.5-2 klx. Self-shading would occur when the trichomes
clump together to form dense mats although the cells at the surface would not be protected. This
clumping behaviour occurs in most motile Oscillatoria species but it was insignificant in the
experiments performed here.
Regular migrations into sediments by 0. terebriformis have been reported by Castenholz
(1982). The causes were unknown although they did not appear to be entirely light-dependent.
The only possible means by which the species investigated here could undergo vertical
migration would depend upon the deposition of an extremely fine and loose sediment. Such sediments do occur in the Medway system but the development of mats upon these has not been
investigated. Clearly, the nature of the sediment is important to mat formation and will
determine which kinds of organisms will be present. The helical path described by the
trichome surfaces results in a screwing effect which would facilitate movement around small
obstacles but the negative correlation between helix pitch and trichome width has no clear
explanation.
The Oscillatoriaceae have been classified into a confusing number of forms (Geitler, 1932;
Desikachary, 1968) although several attempts to rectify this situation have been made (Drouet,
1968; Rippka et al., 1979; Polderman, 1980). However, there is still no satisfactory division of
the family for ecological work, which necessitates a compromise between the classical system of
Geitler and the system proposed by Rippka eta/. (1979) for the strains in pure culture. The value
of filament diameter in the classification of algae has been questioned by Miller & Hoshaw
(1974) but the results obtained here and those of Golubic & Focke (1979) suggest that these
measurements have considerable use in distinguishing between populations of filamentous
cyanobacteria.
The regular seasonal development of M . lyngbyaceus, which occurred throughout the Medway
system, cannot be attributed to any single factor but was probably due to a combination of the
following : increased competition from eukaryotic algae, especially Nitzschia sigma and
Spirogyra nitida; increasing grazing pressure from gastropods during summer, and decreased
sediment disturbance during summer caused by lower rainfall and greater soil cover. Nutrient
limitation was ruled out since the water was continuously eutrophic. The flotation and downstream loss of large areas of dense mat also occurred and this must reduce the likelihood of any
long-term establishment of mats in these sites.
REFERENCES
CASTENHOLZ,
R. W. (1982). Motility and taxes. In The
Biology qf Cyanobacterirj, pp. 414-439. Edited by
N . G. Carr & B. A. Whitton. Oxford: Blackwell
Scientific Publications.
CHU, S. P. (1942). The influence of mineral composition of the medium on the growth of planktonic
algae. Journal of Ecology 30, 284-325.
T. V. (1968). Cyanophyta. New Delhi:
DESIKACHARY,
Indian Council of Agricultural Research.
DROUET,F. (1968). Reiision o f the class~ficationyf' the
Oscillatoriaceae. Monograph Academji uf' Natural
Sciences of Philadelphia 15, 1-370.
FUHS,G . W. (1973). Cytological examination of blue-
green algae. In The Biology of'Blue-green Algae, pp.
117-143. Edited by N . G . Carr & B. A. Whitton.
Oxford: Blackwell Scientific Publications.
GEITLER,L. (1932). C.ranophyceae. In Rabenhorst's
Krjptogamenjlora run Deutschland, Ustrrreich und der
Schweiz, vol. 14. Edited by R. Kolkwitz. Leipzig,
Germany : Akademische Verlagsgesellschaft.
GOLUBIC,
S. (1973). The relationship between bluegreen algae and carbonate deposits. In The Biologj o f
Blue-green Algae, pp. 434-472. Edited by N . G . Carr
& B. A . Whitton. Oxford: Blackwell Scientific Publications.
GOLUBIC,
S. & FOCKE,J . W. (1979). Phormidium
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Mon, 31 Jul 2017 14:13:55
990
A . PENTECOST
hendersonii Howe: identity and significance of a
modern stromatolite building microorganism. Journal of Sedimentary Petrology 48, 75 1-764.
MACKERETH,
F. J. H., HERON,J. & TALLING,J . F.
(1978). Water analysis, some revised methods for
limnologists. Freshwater Biological Association Scientlfic Publication no. 36.
MILLER,R. D. & HOSHAW,R. W . (1974). Cell width as
a taxonomic character with special reference to
Zygnema circumcarinatum Czurda. British Phycological Journal 9, 145-148.
NULTSCH,W. (1962). Der Einfluss des Lichtes auf die
Bewegung der Cyanophyceen. 11. Photokinesis bei
Phormidium autumnale. Planta 57, 61 3-623.
NULTSCH,W. & HELLMAN,
W . (1972). Untersuchungen
zur Photokinesis von Anabaena variabilis Kutzing.
Archic fur Mikrobiologie 82, 76-90.
OLSON,F. C. W. (1950). Quantitative estimates of filamentous algae. Transactions of the American Microscopical Society 59, 272-279.
PENTECOST,
A. (1978). Blue-green algae and freshwater
carbonate deposits. Proceedings of the Royal Society
of London B200, 43-61.
POLDERMAN,
P. J . G. (1980). A key to the Oscillatoriaceae in Northwest European saltmarshes. Blumea
26, 427-437.
RIPPKA,R., DERUELLES,
J.,WATERBURY,
J . B., HERDMAN, M. & STANIER, R. Y. (1979). Generic assignments, strain histories and properties of pure
cultures of cyanobacteria. Journal of General Microbiology 111, 1-61.
ROUND, F. E. & HEATON,J . W. (1966). Persistent,
vertical migration rhythms in benthic microflora.
Journal of Ecology 54, 609-61 5 .
SCHINDLER,
D. W . , SCHMIDT,R. V . & REID, R. A .
(1972). Acidification and bubbling as an alternative
to filtration in determining phytoplankton production by the ‘“C method. Journal of the Fisheries
Research Board of Canada 29, 1627-1 63 1.
WALTER,M. R . (1976). Stromatolites. Devefoprnents in
Sedimentology vol. 20.
ZIMMERMANN,
U. (1969). okologische und physiologische Untersuchungen an der planktischen
Blaualge Oscillatoria rubescens D. C . unter besonderer Berucksichtigung von Licht und Temperatur.
Schweizerische Zeitschrft fur Hydrologie 31, 1-58.
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